Modulation of Clr-b Ligand for the Inhibitory NKR-P1B NK Cell Receptor in Response to Transformation and Genotoxic Stress

Innate immunity is an important first-line defense against infectious disease and malignancies. Natural killer (NK) cells are large granular lymphocytes that are critical for the innate immune response. NK cells recognize target cells undergoing pathological alterations such as viral infection and transformation, through germline-encoded receptors. The recognition of targets undergoing genotoxic stress or transformation by NK cells has been limited to the upregulation of stimulatory NKG2DL and “missing-self” recognition of MHC-I. The ligand pair NKR-P1B:Clr-b participates in “missing-self” recognition of tumour and infected cells, but the role of this axis in sensing stressed cells remains unknown. This thesis focuses on the modulation of Clr-b by genotoxic stress and transformation In the first data chapter, we show that Clr-b is downregulated at the transcript and protein level upon genotoxic stress, in a time- and dose-dependent manner in tumour cells and fibroblasts. This loss occurs upon exposure to a variety of genotoxic agents with the exception of γ-irradiation and radiomimetics. The loss of surface Clr-b results in disinhibition of target cells using 51Cr-release assays and BWZ reporter assays. ii In the next chapter, we investigate the mechanism(s) by which this phenotype occurs. Genotoxic stress-induced Clr-b downregulation is not modulated entirely by classical DNA damage response effectors including ATM, ATR, Chk1, Chk2 and p53. Furthermore, the loss of transcript can by uncoupled from the loss of surface protein following administration of proteasomal inhibitors. Lastly, inhibition of the autocatalytic hammerhead ribozyme harboured within the 3’UTR of the Clr-b transcript can abrogate genotoxic stress-induced Clr-b downregulation. In the final chapter, we investigate the effect of oncogenic stress mediated through H-RasV12 on Clr-b expression in NIH3T3 fibroblasts. H-RasV12-mediated Clr-b downregulation occurs through the MEK-Erk pathway and can be inhibited through pharmacological inhibition of MEK1/2 or shRNA knockdown of Erk2. However, Erk2 alone is not sufficient to induce Clr-b downregulation and requires activation from upstream signals. All together, our results explore the role of genotoxic stress and transformation on the modulation of Clr-b, ligand for the NKR-P1B. This work begins to shed light on the role of NKR-P1B:Clr-b system in “missing-self” recognition of target cells undergoing pathological alterations.

Department dc:contributor.department

Immunology

Year dc:date.issued

2014

Author dc:creator

• Fine, Jason

Advisor dc:contributor.advisor

• Carlyle, James

• NK cell
• genotoxic stress
• transformation
• missing-self recognition

Language dc:language.iso

en_ca

Handle dc:identifier.uri

http://hdl.handle.net/1807/67268

OAI identifier oai:identifier

oai:utoronto.scholaris.ca:1807/67268